In response to signals of either endogenous or exogenous origin, mammalian cells activate a series of events leading to changes in gene expression. In turn, these alterations in gene expression directly participate in the implementation of a global cellular response. While the transcriptional events regulating such changes in gene expression have been thoroughly studied, it is becoming increasingly apparent that posttranscriptional events, which are less well understood, are also major regulatory mechanisms that profoundly modify gene expression. Posttranscriptional events of gene regulation include mRNA processing, transport, stability and translation, as well as protein processing, phosphorylation and degradation. With respect to mRNA stability, we are investigating the mechanisms regulating the expression of various cell cycle regulatory and proliferation-associated genes. We have shown that the expression of cyclins A and B1 throughout the cell division cycle was regulated through the cyclic association of their respective mRNAs with the RNA-binding protein HuR, which results in transcript stabilization during the S phase. We have also reported that high levels of HuR expression in young human fibroblasts contributes to the heightened presence of cyclins A and B1, as HuR stabilizes their mRNAs. Conversely, in senescent human fibroblasts, lower HuR levels directly cause a reduction in the stability, and hence the expression of cyclin A and cyclin B1 mRNAs. Other studies from our laboratory have shown that decreased expression of cyclin D1 following treatment with the stress agent prostaglandin A2 was accomplished through cyclin D1 mRNA destabilization and likely involved the RNA-binding protein AUF1. We previously demonstrated that induction of the cyclin-dependent kinase inhibitor p21 by ultraviolet light and other stresses occurs through stabilization of its mRNA by HuR. Over the past year, we reported that the localization of HuR within the cell is modulated by the AMP-activated kinase (AMPK). This regulatory event is critical for HuR function, as its mRNA stabilizing influence occurs in the cytoplasm. More recently, we have undertaken a high-throughput approach to assess the relative roles of transcription and mRNA turnover in governing gene expression during the cellular stress response. Using cDNA array technology, we have developed a novel approach which involves comparison of large-scale hybridization patterns generated with steady-state mRNA versus newly transcribed (nuclear run-on) RNA. This methodology has allowed us to demonstrate the importance of mRNA turnover in regulating gene expression following several conditions of stress. Our long-term efforts are thus focused on searching for RNA-binding proteins, target mRNA regions, and signaling pathways involved in regulating the stability of mRNAs encoding growth control and cell cycle regulatory genes. The product of the tumor suppressor gene von Hippel- Lindau (pVHL) is believed to modulate gene expression at the levels of transcription elongation, mRNA stability and protein degradation. Our investigation of pVHL's global influence on gene expression using SAGE was recently reported. Now, we have focused our efforts towards understanding VHL?s influence on protein translation. We anticipate that these studies will help elucidate pVHL's function and its tumor suppressor properties.